The need for an effective vaccine against HlV-1 is as urgent as ever. It is currently widely accepted that for a vaccine against HIV to be effective, it should elicit broad and potent anti-HIV neutralizing antibody responses. Anti-HIV neutralizing antibodies (NAbs) target the HIV envelope glycoprotein (Env) on the surface of infectious virions. Despite recent advances in the design of HIV Env-based immunogens, such immunogens have not yet elicited broadly cross-reactive NAbs against primary HIV viruses. The primary reason for this, is that conserved neutralization epitopes, which are present and exposed on such immunogens, are not immunogenic in the context of vaccination. Improving the immunogenicity of such epitopes is crucial for the elicitation of broad anti-HIV neutralizing antibody responses. We currently lack fundamental knowledge on the way B cells recognize such conserved neutralization epitopes on our immunogens and how they subsequently respond to that recognition. In this Project Three, we will investigate how the binding affinity and the duration of the immunogen-BCR interaction, influences the ensuing intracellular events that lead to antibody production. Our plan is to take a 'step back'and examine in detail the interaction between immunogens that express a single HIV neutralization epitope and B cells that express the BCR of that epitope. We will focus our efforts on the epitope recognized by the broadly neutralizing antibody b12. We will determine how the biophysical properties of this interaction define the nature of the ensuing intracellular signaling events that lead to b12-specific B cell antibody responses. These in vitro studies will be complemented by in vivo studies in rhesus macaques (Core B), during which the fate of b12 BCR-expressing B cells will be monitored in the context of immunization and infection. These macaque studies will inform on the evolution of the b12 B cell responses in the contexts of a temporal presentation of immunogen (immunization) and during chronic presentation of the immunogen (SHIV-infection) and on how b12 antibody affinity maturation influences viral-evolution and vice versa.